Regulation and evaluation of five methanol-inducible promoters in the methylotrophic yeast Candida boidinii

We isolated the promoter regions of five methanol-inducible genes (P-AOD1, alcohol oxidase: P-DAS1, dihydroxyacetone synthase; P-FDll1, formate dehydrogenase; P-PMP20, Pmp20; and P-PMP47, Pmp47) from the Candida boidinii genome, and evaluated their strength and studied their regulation using the acid phosphatase gene of Saccharomyces cerevisiae (ScPHO5) as the reporter. Of the five promoters, P-DAS1 was the strongest methanol-inducible promoter whose strength was approximately 1.5 times higher than that of the commonly used P-AOD1 in methanol-induced cells. Although the expression of P-AOD1 and P-DAS1 was completely repressed by the presence of glucose, formate-induced expression of P-FDH1 was not repressed by glucose. Expression under P-PMP47, another methanol-inducible promoter, was highly induced by oleate. The induction kinetics of P-PMP47 and P-DAS1 revealed that methanol induces the expression of peroxisome membrane protein Pmp47, earlier than the expression of matrix enzyme dihydroxyacetone synthase (Das1p), and that this information is contained in the promoter region of the respective gene. This is the first report which evaluates several methanol-inducible promoters in parallel in the methylotrophic yeast. (C) 2000 Elsevier Science B.V. All rights reserved.